Human granulocytic ehrlichiosis (HGE) is a new tick-borne zoonosis that is increasingly recognized as a threat to public health in the United States. HGE is a systemic febrile illness often accompanied by hematological abnormalities including leukopenia and thrombocytopenia. It frequently requires prolonged hospitalization and when the treatment is delayed due to misdiagnosis or in immunocompromised patients, HGE can be fatal. HGE is caused by infection of peripheral blood granulocytes with an obligate intracellular bacterium, an ehrlichia sp. called an HGE agent. The investigators previously showed that a family of 44-kDa-range major outer membrane proteins of the HGE agent is the immunodominant antigen in human infection. They demonstrated that 44-kDa proteins (P44s) are encoded by a multigene family. Their recent study indicates that there are a total of approximately 18 copies of p44 genes and the HGE agent in a human promyelocytic leukemia cell line, HL-60, expresses 5 of them. These p44 genes are expressed at different sites that are widely distributed throughout the genome, suggesting a potential unique mechanism of providing antigenic diversity in the HGE agent. This system of P44 major surface proteins may provide an instructive model for studying adaptive genetic strategies, and fundamental surface properties, of several ehrlichial species contributing to human and animal diseases. Structural and combinatorial variation in P44 profiles expressed on the unique intracellular bacterial surface, may profoundly affect in-host adaptive capabilities of ehrlichiae. Their specific aims will address important questions in the understanding of this prototype system, including: (1) the degree, pattern and variation of p44 genes expressed by the HGE agent in its different hosts (tick vector, rodent reservoir, and accidental host), (2) the range of variation in the natural repertoire of p44 genes and expression among HGE agent strains, (3) the antibody response to each P44 protein in infected animals and humans, and (4) P44 protein interactions and p44 gene expression mechanisms. Approaches include RT-PCR and sequencing-based characterization of p44 genes expressed by HGE agents in experimentally infected animals and ticks and in several human isolates, cloning the p44 genes expressed, analysis of antibody responses to individual P44-specific peptides, analysis of P44 complexes and a unique mRNA splicing mechanism. These efforts are expected to reveal an important role for the p44-multigene family in strategies of adaptive surface variation and avoidance of host immune responses.